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Submitted on December 23, 2002
Accepted on May 13, 2003
1 Signalisation cellulaire, Régulation de gènes et Physiologie de l'Axe gonadotrope, Centre National de la Recherche Scientifique-UMR 7079, Physiologie et Physiopathologie, Université Pierre et Marie Curie, 75252 Paris, France
* To whom correspondence should be addressed. E-mail: raymond.counis{at}snv.jussieu.fr.
The neuronal nitric oxide synthase (NOS I) is expressed and hormonally regulated in rat anterior pituitary gonadotrophs. In the present study, we investigated the mechanisms that underlie the constitutive and GnRH up-regulated activity of the pituitary exon 1p promoter of the NOS I gene in these cells. Through the use of 5'-deletions and transient transfections in L
T2, a gonadotrope-derived cell line, we delineated a NOS I cell-specific (NCS) enhancer region (-73/-59) that is required for constitutive activity. Independently of the NCS enhancer, GnRH responsiveness is supported by a bipartite regulatory domain referred to as the GnRH response element (GnRE) I and II located between -33/-10 and -4/+4, the latter consisting of a cAMP-like response element. By combining transient transfections, gel shift and supershift assays we demonstrate that Sp1 and LIM-homeodomain-related protein bind the NCS enhancer whereas CREB and CREM-like factors bind the GnRE II motif. We further show that factors involved in GnRH regulation are also implicated in constitutive activity suggesting intimate links between constitutive and regulated promoter activity. We speculate that specific expression of the NOS I gene in gonadotrophs together with its regulation by GnRH is suggestive of a critical participation of NOS I in gonadotrope function.
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